Amber Building Glycoprotein Tutorial

Glycoproteins are proteins which contain oligosaccharide chains (glycans) covalently attached to amino acid side-chains. The carbohydrate is attached to the protein in a cotranslational or posttranslational modification. This process is known as glycosylation. Secreted extracellular proteins are often glycosylated. Carbohydrates are attached to some proteins to form glycoproteins. Glycoproteins are also often important integral membrane proteins, where they play a role in cell–cell interactions.

The following content is summarized from the Amber 16 manual.

Tutorials

Use LEAP to construct glycoprotein

The LEAP command we give is based on the premise that you already have a PDB file containing sugar molecules and proteins, and the relative conformation between the two is appropriate. Even in order to connect the simplest sugar with the simplest protein, you are required Have a thorough understanding of LEAP commands.

Set the PDB file of glycosylated protein in LEAP

1. Delete all HETATM atoms in the PDB file. These atoms usually include bound ligands, amorphous water molecules and non-coordinating metal ions. If there are hydrogen atoms, delete them all.

2. Check the beginning of the PDB file to see if there is a disulfide bond (SSBOND). The name of the cysteine residue needs to be changed from CYS to CYX.

3. Prepare a PDB file containing protein and sugar molecules, where the sugar molecules are oriented appropriately relative to the protein surface. Methods include:

✓ Use a molecular modeling program to align the sugar molecules generated by GLYCAM with the protein, and save the coordinates in a single file. Remember to delete the OH or OMe at the end of the sugar molecule.

✓ Use the online glycoprotein construction tool http://www.glycam.org. This tool allows users to upload the coordinates of the protein, construct the sugar molecule (or select from the library), and connect it to the protein. Then you can download the desired AMBER file.

Script

Enter the following command in xleap (if you don't need a graphical interface, you can use tleap) or save the command to a file and source.

source leaprc.GLYCAM_06j-1 # load the GLYCAM-06 leaprc for ff14SB

source leaprc.protein.ff14SB # load the protein force field

glyprot = loadpdb 3rn3_nlink.pdb # load protein and glycan pdb file

bond glyprot.125.O4 glyprot.126.C1 # make inter glycan bonds

bond glyprot.126.O4 glyprot.127.C1

bond glyprot.127.O6 glyprot.128.C1

bond glyprot.127.O3 glyprot.129.C1

bond glyprot.34.SG glyprot.125.C1 # make glycan -- protein bond

bond glyprot.26.SG glyprot.84.SG # make disulfide bonds

bond glyprot.40.SG glyprot.95.SG

bond glyprot.58.SG glyprot.110.SG

bond glyprot.65.SG glyprot.72.SG

addions glyprot CL 0 # neutralize appropriately

solvateBox glyprot TIP3P BOX 8 # solvate the solute

savepdb glyprot 3nr3_glycan.pdb # save pdb file

saveamberparm glyprot 3nr3_glycan.top 3nr3_glycan.crd # save top, crd

quit # exit leap

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